There is evidence for cross talk between the immune system and the equally complex and important nervous system. Immune system status is clearly influenced by status of the nervous system, and the opposite is also true. Studies by participants in this project have added to an evolving literature suggesting that the natural chemical neurotransmitter, acetylcholine (ACh), and the tobacco alkaloid, nicotine, have physiologically-relevant effects on immune system development and function. The diverse family of nicotinic acetylcholine receptors (nAChR) are targets for ACh and nicotine signaling. Studies by participants in this project indicate that specific nAChR subtypes are expressed by cells of the immune system, suggesting that these nAChR mediate effects of ACh and nicotine on immune system development and function. Based on these findings, this multi-investigator project will test the central hypothesis that nicotine and related ligands can regulate immune system development and function. The project will establish effects of nicotinic ligands on several important parameters relating to immune system function and development with a particular focus on T cell maturation. Work in the project will also identify and characterize known and/or potentially novel nAChR. Immune system components and thymic organ cultures from humans and mice will be used to characterize effects of nicotinic drug exposures on numbers and proportions of phenotypically-distinct T cell subsets and their expression of markers for positive/negative selection. Other studies will identify immune system nAChR subunits and subtypes and the cell subsets that express them with reference to developmental and drug treatment effects. Assays for cytokine production and for cytokine or recombinase activating gene expression using organ or cell culture samples will be conducted to identify mechanisms by which nicotine and related compounds affect immune cell development. Hypotheses to be addressed include: I) Nicotine regulates the development of T cells in the thymus. 2) Specific nicotinic ligands block or mimic nicotine's effects and reveal influences of endogenous signaling by ACh on T cell development. 3) nAChR involved in immune system development are expressed by T cells and their progenitors and/or by thymic stromal cells in developmentally-relevant patterns. 4) Immune system nAChR mediate their effects on T cell development by altering expression and/or secretion of cytokines and/or by altering expression of genes involved in T cell receptor rearrangements. One of the biomedically-relevant hypotheses of this project is that tobacco use and nicotine exposure affect immune system development.